scholarly journals Multicenter Evaluation of BD Max Enteric Parasite Real-Time PCR Assay for Detection of Giardia duodenalis, Cryptosporidium hominis, Cryptosporidium parvum, and Entamoeba histolytica

2016 ◽  
Vol 54 (11) ◽  
pp. 2681-2688 ◽  
Author(s):  
S. Madison-Antenucci ◽  
R. F. Relich ◽  
L. Doyle ◽  
N. Espina ◽  
D. Fuller ◽  
...  
Keyword(s):  
Entamoeba Histolytica ◽  
Cryptosporidium Parvum ◽  
Fluorescent Antibody ◽  
Giardia Duodenalis ◽  
The United States ◽  
Parasitic Infections ◽  
Pcr Assay ◽  
Cryptosporidium Hominis ◽  
Content Type ◽  
Common Causes

Common causes of chronic diarrhea among travelers worldwide include protozoan parasites. The majority of parasitic infections are caused by Giardia duodenalis , Entamoeba histolytica , Cryptosporidium parvum , and Cryptosporidium hominis . Similarly, these species cause the majority of parasitic diarrhea acquired in the United States. Detection of parasites by gold standard microscopic methods is time-consuming and requires considerable expertise; enzyme immunoassays and direct fluorescent-antibody (DFA) stains have lowered hands-on time for testing, but improvements in sensitivity and technical time may be possible with a PCR assay. We performed a clinical evaluation of a multiplex PCR panel, the enteric parasite panel (EPP), for the detection of these common parasites using the BD Max instrument, which performs automated extraction and amplification. A total of 2,495 compliant specimens were enrolled, including 2,104 (84%) specimens collected prospectively and 391 (16%) specimens collected retrospectively. Approximately equal numbers were received in 10% formalin (1,273 specimens) and unpreserved (1,222 specimens). The results from the EPP were compared to those from alternate PCR and bidirectional sequencing (APCR), as well as DFA ( G. duodenalis and C. parvum or C. hominis ) or trichrome stain ( E. histolytica ). The sensitivity and specificity for prospective and retrospective specimens combined were 98.2% and 99.5% for G. duodenalis , 95.5% and 99.6 for C. parvum or C. hominis , and 100% and 100% for E. histolytica , respectively. The performance of the FDA-approved BD Max EPP compared well to the reference methods and may be an appropriate substitute for microscopic examination or immunoassays.

scholarly journals Comparison of Assays for Sensitive and Reproducible Detection of Cell Culture-Infectious Cryptosporidium parvum and Cryptosporidium hominis in Drinking Water

2011 ◽  
Vol 78 (1) ◽  
pp. 156-162 ◽  
Author(s):  
Anne M. Johnson ◽  
George D. Di Giovanni ◽  
Paul A. Rochelle
Keyword(s):  
Cell Culture ◽  
Drinking Water ◽  
Cryptosporidium Parvum ◽  
False Positives ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Cryptosporidium Hominis ◽  
Content Type ◽  
Detection Assay ◽  
Pcr Targeting

ABSTRACTThis study compared the three most commonly used assays for detectingCryptosporidiumsp. infections in cell culture: immunofluorescent antibody and microscopy assay (IFA), PCR targetingCryptosporidiumsp.-specific DNA, and reverse transcriptase PCR (RT-PCR) targetingCryptosporidiumsp.-specific mRNA. Monolayers of HCT-8 cells, grown in 8-well chamber slides or 96-well plates, were inoculated with a variety of viable and inactivated oocysts to assess assay performance. All assays detected infection with low doses of flow cytometry-enumeratedCryptosporidium parvumoocysts, including infection with one oocyst and three oocysts. All methods also detected infection withCryptosporidium hominis. The RT-PCR assay, IFA, and PCR assay detected infection in 23%, 25%, and 51% of monolayers inoculated with threeC. parvumoocysts and 10%, 9%, and 16% of monolayers inoculated with one oocyst, respectively. The PCR assay was the most sensitive, but it had the highest frequency of false positives with mock-infected cells and inactivated oocysts. IFA was the only infection detection assay that did not produce false positives with mock-infected monolayers. IFA was also the only assay that detected infections in all experiments with spiked oocysts recovered from Envirochek capsules following filtration of 1,000 liters of treated water. Consequently, cell culture with IFA detection is the most appropriate method for routine and sensitive detection of infectiousCryptosporidium parvumandCryptosporidium hominisin drinking water.

scholarly journals Performance and Verification of a Real-Time PCR Assay Targeting thegyrAGene for Prediction of Ciprofloxacin Resistance in Neisseria gonorrhoeae: TABLE 1

2016 ◽  
Vol 54 (3) ◽  
pp. 805-808 ◽  
Author(s):  
P. Hemarajata ◽  
S. Yang ◽  
O. O. Soge ◽  
R. M. Humphries ◽  
J. D. Klausner
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Real Time ◽  
Real Time Pcr ◽  
The United States ◽  
Test Results ◽  
Pcr Assay ◽  
Content Type ◽  
Agar Dilution ◽  
Ciprofloxacin Resistance ◽  
To Receive

In the United States, 19.2% ofNeisseria gonorrhoeaeisolates are resistant to ciprofloxacin. We evaluated a real-time PCR assay to predict ciprofloxacin susceptibility using residual DNA from the Roche Cobas 4800 CT/NG assay. The results of the assay were 100% concordant with agar dilution susceptibility test results for 100 clinical isolates. Among 76 clinical urine and swab specimens positive forN. gonorrhoeaeby the Cobas assay, 71% could be genotyped. The test took 1.5 h to perform, allowing the physician to receive results in time to make informed clinical decisions.

scholarly journals Comparison of Automated Quantitative Reverse Transcription-PCR and Direct Fluorescent-Antibody Detection for Routine Rabies Diagnosis in the United States

2015 ◽  
Vol 53 (9) ◽  
pp. 2983-2989 ◽  
Author(s):  
Michelle Dupuis ◽  
Scott Brunt ◽  
Kim Appler ◽  
April Davis ◽  
Robert Rudd
Keyword(s):  
United States ◽  
Reverse Transcription ◽  
Gold Standard ◽  
Fluorescent Antibody ◽  
The United States ◽  
Pcr Assay ◽  
Qrt Pcr ◽  
Reverse Transcription Pcr ◽  
Direct Fluorescent Antibody ◽  
Rabies Diagnosis

Rabies virus found worldwide and prevalent throughout the United States continues to be a public health concern. Direct-fluorescent antibody (DFA) detection remains the gold standard for rabies virus diagnostics. Assessing the utility of a high-throughput molecular platform such as the QIAsymphony SP/AS, in conjunction with quantitative reverse transcription-PCR (qRT-PCR), to augment or potentially replace the DFA test, was the focus of this project. Here we describe a triplex qRT-PCR assay, including assembly and evaluation for sensitivity, specificity, and ability to detect variants. Additionally, we compared the qRT-PCR assay to the gold standard direct fluorescent-antibody test. More than 1,000 specimens submitted for routine rabies diagnosis were tested to directly compare the two methods. All results were in agreement between the two methods, with one additional specimen detected by qRT-PCR below the limits of the DFA sensitivity. With the proper continued validation for variant detection, molecular methods have a place in routine rabies diagnostics within the United States.

Fate of Cryptosporidium parvum and Cryptosporidium hominis oocysts and Giardia duodenalis cysts during secondary wastewater treatments

2009 ◽  
Vol 105 (3) ◽  
pp. 689-696 ◽  
Author(s):  
Hui-Wen A. Cheng ◽  
Frances E. Lucy ◽  
Thaddeus K. Graczyk ◽  
Michael. A. Broaders ◽  
Leena Tamang ◽  
...  
Keyword(s):  
Cryptosporidium Parvum ◽  
Giardia Duodenalis ◽  
Cryptosporidium Hominis

scholarly journals Evaluation of Level of Agreement in Bordetella Species Identification in Three U.S. Laboratories during a Period of Increased Pertussis

2015 ◽  
Vol 53 (6) ◽  
pp. 1842-1847 ◽  
Author(s):  
Brunilís Burgos-Rivera ◽  
Adria D. Lee ◽  
Katherine E. Bowden ◽  
Amanda E. Faulkner ◽  
Brent L. Seaton ◽  
...  
Keyword(s):  
Species Identification ◽  
Insertion Sequence ◽  
The United States ◽  
Pcr Assay ◽  
Assay Sensitivity ◽  
Content Type ◽  
Negative Results ◽  
Relative Contribution ◽  
Level Of Agreement ◽  
Good Agreement

While PCR is the most common method used for detectingBordetella pertussisin the United States, most laboratories use insertion sequence481(IS481), which is not specific forB. pertussis; therefore, the relative contribution of otherBordetellaspecies is not understood. The objectives of this study were to evaluate the proportion of otherBordetellaspecies misidentified asB. pertussisduring a period of increased pertussis incidence, determine the level of agreement inBordetellaspecies detection between U.S. commercial laboratories and the CDC, and assess the relative diagnostic sensitivity of CDC's PCR assay when using a different PCR master mix. Specimens collected between May 2012 and May 2013 were tested at two U.S. commercial laboratories forB. pertussisandB. parapertussisdetection. Every fifth specimen positive for IS481and/or IS1001with cycle threshold (CT) values of ≤35 was sent to CDC for PCR testing that identifiesBordetellaspecies. Specimens with indeterminate or negative results in the CDC PCR were tested using an alternate PCR master mix. Of 755 specimens, there was agreement in species identification for 83.4% (n= 630). Of the specimens with different identifications (n= 125), 79.2% (n= 99) were identified as indeterminateB. pertussisat CDC. Overall, 0.66% (n= 5) of the specimens were identified asB. holmesiiorB. bronchisepticaat CDC. Of 115 specimens with indeterminate or negative results, 46.1% (n= 53) wereB. pertussispositive when tested by an alternate master mix, suggesting a possible increase in assay sensitivity. This study demonstrates good agreement between the two U.S. commercial laboratories and CDC and little misidentification ofBordetellaspecies during the 2012 U.S. epidemic.

Prevalence of intestinal parasitic infections among diarrheic patients from Ahvaz, southwestern Iran

2020 ◽  
Vol 20 ◽  
Author(s):  
Saina Karami ◽  
Molouk Beiromvand ◽  
Kobra Kohansal
Keyword(s):  
Entamoeba Histolytica ◽  
Intestinal Parasites ◽  
Giardia Duodenalis ◽  
Gastrointestinal Parasites ◽  
Parasitic Infections ◽  
Blastocystis Hominis ◽  
Cross Sectional ◽  
Intestinal Parasitic Infections ◽  
Cryptosporidium Spp ◽  
Southwestern Iran

Background:: Intestinal parasitic infections are one of the most common infections in humans, particularly in re-source-poor communities. Gastrointestinal parasites, specially protozoa can lead to diarrhea, malabsorption, and anemia. The majority of parasitic diarrhea is caused by Entamoeba histolytica, Giardia duodenalis, and Cryptosporidium spp.. The present study aimed to evaluate the prevalence of intestinal parasites among diarrheic patients referred to the Shahid Rajaee Polyclinic, Ahvaz, southwestern Iran. Methods:: A cross-sectional study was conducted to determine the prevalence of intestinal parasites among 250 diarrheic pa-tients using direct smear, formalin-ether concentration, ziehl- neelsen, and trichrome staining. Results:: The results indicated that 34.4% (86/250) of the patients were infected with pathogenic parasites. Giardia duode-nalis with an occurrence of 18.8% (47/250) and Cryptosporidium spp. with a frequency of 2.8% (7/250) had the highest and lowest infection rates, respectively. Blastocystis hominis with a frequency of 15.2% (38/250) showed the highest prevalence rate after G. duodenalis. Entamoeba histolytica/dispar was observed in 3 (1.2%) of diarrheic patients. The age group 1−10 years old was the most frequently infected group (27.9%). We could not find a significant association between the source of drinking water and intestinal parasitic infections (p= 0.912). Conclusions:: This study demonstrated that G. duodenalis was the predominant parasite found among the patients. The re-sults revealed that intestinal parasites were one of the main health problems in the region.

scholarly journals Drug Repurposing Screen Reveals FDA-Approved Inhibitors of Human HMG-CoA Reductase and Isoprenoid Synthesis That Block Cryptosporidium parvum Growth

2013 ◽  
Vol 57 (4) ◽  
pp. 1804-1814 ◽  
Author(s):  
Kovi Bessoff ◽  
Adam Sateriale ◽  
K. Kyungae Lee ◽  
Christopher D. Huston
Keyword(s):  
Cryptosporidium Parvum ◽  
Financial Incentives ◽  
Diarrheal Disease ◽  
Drug Repurposing ◽  
Bioinformatic Analysis ◽  
Current Standard ◽  
Cryptosporidium Hominis ◽  
Hmg Coa Reductase ◽  
Content Type ◽  
Coa Reductase

ABSTRACTCryptosporidiosis, a diarrheal disease usually caused byCryptosporidium parvumorCryptosporidium hominisin humans, can result in fulminant diarrhea and death in AIDS patients and chronic infection and stunting in children. Nitazoxanide, the current standard of care, has limited efficacy in children and is no more effective than placebo in patients with advanced AIDS. Unfortunately, the lack of financial incentives and the technical difficulties associated with working withCryptosporidiumparasites have crippled efforts to develop effective treatments. In order to address these obstacles, we developed and validated (Z′ score = 0.21 to 0.47) a cell-based high-throughput assay and screened a library of drug repurposing candidates (the NIH Clinical Collections), with the hopes of identifying safe, FDA-approved drugs to treat cryptosporidiosis. Our screen yielded 21 compounds with confirmed activity againstC. parvumgrowth at concentrations of <10 μM, many of which had well-defined mechanisms of action, making them useful tools to study basic biology in addition to being potential therapeutics. Additional work, including structure-activity relationship studies, identified the human 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase inhibitor itavastatin as a potent inhibitor ofC. parvumgrowth (50% inhibitory concentration [IC50] = 0.62 μM). Bioinformatic analysis of theCryptosporidiumgenomes indicated that the parasites lack all known enzymes required for the synthesis of isoprenoid precursors. Additionally, itavastatin-induced growth inhibition ofC. parvumwas partially reversed by the addition of exogenous isopentenyl pyrophosphate, suggesting that itavastatin reducesCryptosporidiumgrowth via on-target inhibition of host HMG-CoA reductase and that the parasite is dependent on the host cell for synthesis of isoprenoid precursors.

scholarly journals Multicenter Clinical Evaluation of the Automated Aries Group A Strep PCR Assay from Throat Swabs

2018 ◽  
Vol 57 (2) ◽  
Author(s):  
N. Kanwar ◽  
J. Crawford ◽  
C. Ulen ◽  
T. S. Uphoff ◽  
J. Dien Bard ◽  
...  
Keyword(s):  
Predictive Value ◽  
Throat Swab ◽  
False Negative ◽  
Reference Method ◽  
The United States ◽  
Antibiotic Administration ◽  
Pcr Assay ◽  
Assay Sensitivity ◽  
Content Type ◽  
Group A

ABSTRACT Group A Streptococcus (GAS) is one of the leading causes of bacterial pharyngitis. Early GAS diagnosis is critical for appropriate antibiotic administration that reduces the risk of GAS sequelae and limits spread of the infection. The Aries Group A Strep (GAS) assay (Luminex, Austin, TX) is a fully automated PCR assay for direct detection of GAS in throat swab specimens in less than 2 h with minimum hands-on time. This multicenter prospective study evaluated the clinical performance of the Aries GAS assay compared to that of Streptococcus pyogenes culture. Subjects with symptoms consistent with pharyngitis were enrolled across four sites in the United States, and a throat swab in liquid Amies medium was obtained. Aries and reference testing was performed within 72 and 48 h after sample collection, respectively. Of 623 throat swab specimens from patients with pharyngitis (93.6% <18 years old, 54.3% female), the reference method yielded valid results for 618 specimens. Reference and Aries assay testing showed GAS-positive results for 160 (25.9%) and 166 (26.9%) specimens, respectively. Compared to the reference method, Aries assay sensitivity was 97.5% (95% confidence interval [CI], 93.7% to 99.0%), specificity was 97.8% (95% CI, 96.0 to 98.8%), positive predictive value was 94.0% (95% CI, 89.3% to 96.7%), and negative predictive value was 99.1% (95% CI, 97.7% to 99.7%). There were 10 false-positive and four false-negative detections with the Aries assay. Discrepant analysis with bidirectional sequencing yielded concordant results with the Aries assay for nine of 14 discordant samples. The Aries assay had high sensitivity and specificity for qualitative detection of group A Streptococcus from patients with pharyngitis.

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